Original Article
DOI: 10.1590/0103-0582201432214813
Respiratory infections in children up to two years of age on prophylaxis with palivizumab Infecções respiratórias em crianças menores de dois anos de idade submetidas a profilaxia com palivizumabe Infecciones respiratorias en niños menores de dos años de edad sometidos a profilaxis con palivizumabe Ana Isabel M. P. Monteiro1, Nancy Cristina J. Bellei1, Alessandra Ramos Sousa1, Amélia Miyashiro N. dos Santos1, Lily Yin Weckx1
ABSTRACT
Objective: To identify the viruses involved in acute respiratory tract infections and to analyze the rates of hospitalization and death in children on palivizumab prophylaxis. Methods: Prospective cohort of 198 infants up to one year old who were born before 29 weeks of gestational age and infants under two years old with hemodynamically unstable cardiopathy or chronic pulmonary disease who received prophylactic palivizumab against severe respiratory syncytial virus infections in 2008. During the study period, in each episode of acute respiratory tract infection, nasopharyngeal aspirate was collected to identify respiratory syncytial virus, adenovirus, parainfluenza 1, 2 and 3, influenza A and B by direct immunofluorescence, rhinovirus and metapneumovirus by polymerase chain reaction preceded by reverse transcription. Data regarding hospitalization and deaths were monitored. Results: Among the 198 studied infants, 117 (59.1%) presented acute respiratory tract infections, with a total of 175 episodes. Of the 76 nasopharyngeal aspirates collected during respiratory tract infections, 37 were positive, as follow: rhinovirus (75.7%), respiratory syncytial virus (18.9%), parainfluenza (8.1%), adenovirus 2 (2.7%), metapneumovirus (2.7%) and three samples presented multiple
Instituição: Escola Paulista de Medicina da Universidade Federal de São Paulo (Unifesp), São Paulo, SP, Brasil 1
Escola Paulista de Medicina da Unifesp, São Paulo, SP, Brasil
agents. Of the 198 children, 48 (24.4%) were hospitalized: 30 (15.2%) for non-infectious etiology and 18 (9.1%) for respiratory causes. Among these 18 children, one case of respiratory syncytial virus was identified. Two deaths were reported, but respiratory syncytial virus was not identified. Conclusions: During the prophylaxis period, low frequency of respiratory syncytial virus infections and low rates of hospitalization were observed, suggesting the benefit of palivizumab prophylaxis. Key-words: respiratory syncytial viruses; antibodies, monoclonal; respiratory tract infections/prevention & control; infant. RESUMO
Objetivo: Identificar os vírus envolvidos nos quadros de infecções agudas de trato respiratório e analisar as taxas de internação e de óbito em crianças submetidas à profilaxia com palivizumabe. Métodos: Coorte prospectiva com 198 crianças menores de um ano nascidas antes de 29 semanas de idade gestacional e crianças menores de dois anos com cardiopatia hemodinamicamente instável ou doença pulmonar crônica que receberam palivizumabe para profilaxia contra infecções graves pelo vírus
Endereço para correspondência: Ana Isabel M. P. Monteiro Rua Loefgreen, 1.998 CEP 04040-003 – São Paulo/SP E-mail: [email protected] Fonte Financiadora: Abbott Laboratories (Chicago, EUA) custeou os testes laboratoriais Conflito de interesse: Lily Yin Weckx realiza pesquisa clínica financiada pela GlaxoSmithKline, Merck Sharp & Dome, Novartis, Sanofi e Pfizer e palestras científicas para a GSK, MSD, Abbott, Sanofi, Novartis e Pfizer. Os demais autores não apresentam conflito de interesse Recebido em: 23/9/2013 Aprovado em: 27/11/2013
Rev Paul Pediatr 2014;32(2):152-8.
Ana Isabel M. P. Monteiro et al
sincicial respiratório, em 2008. No período do estudo, em cada episódio de infecção aguda do trato respiratório, coletou-se aspirado de nasofaringe para identificar vírus sincicial respiratório, adenovírus, parainfluenza 1, 2 e 3, influenza A e B por imunofluorescência direta, rinovírus e metapneumovírus por reação em cadeia de polimerase precedida de transcriptase reversa. Monitoraram-se internações e óbitos nesse grupo. Resultados: Das 198 crianças acompanhadas, 117 (59,1%) apresentaram infecções agudas de trato respiratório, totalizando 175 episódios. De 76 aspirados de nasofaringe coletados na vigência de infecções do trato respiratório, 37 foram positivos, encontrando-se: rinovírus (75,7%), vírus sincicial respiratório (18,9%), parainfluenza (8,1%), adenovírus (2,7%), metapneumovírus (2,7%) e múltiplos agentes em três amostras. Das 198 crianças, 48 (24,4%) foram internadas, sendo 30 (15,2%) por etiologia não respiratória e 18 (9,1%) por problemas respiratórios; entre os 18 casos, um foi por vírus sincicial respiratório. Duas crianças evoluíram para óbito, não tendo sido identificado o vírus sincicial respiratório. Conclusões: Na vigência de profilaxia, observou-se frequência baixa de infecções pelo vírus sincicial respiratório e baixo índice de hospitalizações, sugerindo benefício da profilaxia com palivizumabe. Palavras-chave: vírus sinciciais respiratórios; anticorpos monoclonais; infecções respiratórias/prevenção & controle; lactente. RESUMEN
Objetivo: Identificar los virus implicados en los cuadros de infecciones agudas de trato respiratorio y analizar las tasas de internación y de óbito en niños sometidos a la profilaxis con palivizumabe. Métodos: Cohorte prospectiva con 198 niños con menos de un año nacidas antes de 29 semanas de edad gestacional y niños con menos de dos años con cardiopatía hemodinámicamente inestable o enfermedad pulmonar crónica que recibieron palivizumabe para profilaxis contra infecciones graves por el virus sincitial respiratorio, en 2008. En el periodo de estudio, en cada episodio de infección aguda del trato respiratorio, se recogió aspirado de nasofaringe para identificar virus sincitial respiratorio, adenovirus, parainfluenza 1,2 y 3, influenza A y B por técnica del anticuerpo fluorescente directa, rinovirus y metapneumovirus por reacción en cadena de la polimerasa precedida por transcriptasa inversa. Se monitorean internaciones y óbitos en ese grupo.
Rev Paul Pediatr 2014;32(2):152-8.
Resultados: De los 198 niños seguidos, 117 (59,1%) presentaron infecciones agudas de trato respiratorio, totalizando 175 episodios. De 76 aspirados de nasofaringe recogidos en la vigencia de infecciones de trato respiratorio, 37 fueron positivos, encontrándose: rinovirus (75,7%), virus sincitial respiratorio (18,9%), parainfluenza (8,1%), adenovirus (2,7%), metapneumovirus (2,7%) y múltiples agentes en tres muestras. De los 198 niños, 48 (24,4%) fueron internados, siendo 30 (15,2%) por etiología no respiratoria y 18 (9,1%) por problemas respiratorios; entre los 18 casos, uno fue por virus sincitial respiratorio. Dos niños evolucionaron a óbito, no habiendo sido identificado el virus sincitial respiratorio. Conclusiones: En la vigencia de profilaxis, se observó frecuencia baja de infecciones por el virus sincitial respiratorio y bajo índice de hospitalizaciones, sugiriendo beneficio de la profilaxis con palivizumabe. Palabras clave: virus sincitiales respiratorios; anticuerpos monoclonales; infecciones respiratorias/ prevención y control; lactante.
Introduction Respiratory syncytial virus (RSV) is the major cause of acute lower respiratory tract infections in children younger than 2 years of age. It has a global distribution and a seasonal behavior. It is estimated that, worldwide, 33.8 million new episodes of acute lower respiratory tract infections by RSV occur annually in children under 5 years of age. Among these, about 3.4 million require hospitalization and 66 to 199 thousand children evolve to death, 99% occurring in developing countries(1). In general, primary RSV infection evolves as a common cold. However, approximately 25% of children under 2 years may present, in its first episode, infection of the lower airways, severe respiratory failure, need for ventilatory support, and, occasionally, death(2). Preterm children with chronic pulmonary disease, immunocompromised children, and children with heart diseases (particularly cyanotic) present greater risk of severe respiratory disease by RSV(3-6). Some environmental factors are also associated with higher incidence of RSV infections, such as exposure to smoke, attendance at day care centers, or household contact with other children(4,5). In tropical and subtropical countries, the occurrence of RSV infections is greater in fall and winter, with significant regional variations(7). Epidemiological studies on RSV in the Southeast region of Brazil showed that the infection by RSV starts in April, with peaks in May and June(2,8). More recent
153
Respiratory infections in children up to two years of age on prophylaxis with palivizumab
studies conducted in the municipality of São Paulo showed that the RSV season occurs from March to July, and it may start earlier or last up until August(9,10). There is no specific treatment for RSV infection(7,11), and prophylaxis with a specific anti-RSV monoclonal antibody (palivizumab) is widely recommended for high-risk cases(4-6). Its use demonstrated a reduction in the rates of hospitalization by RSV infection (55%), hospitalization in ICUs (57%), length of hospital stay (42%), and the number of days with need of oxygen by 40%(4,5,12,13). Palivizumab is, however, a high cost medication. In Brazil, until 2007, many children received this medication after judicial demands, not always based in technical criteria(14). The resolution SS n. 249, from July 13, 2007, from the Health Secretariat of the State of São Paulo(15), made palivizumab freely available for SUS (Brazilian public Unified Health System) patients who were classified as “highly recommended” by the Brazilian Pediatrics Society(5). Some centers were indicated by the State Secretariat as a place for the application of the palivizumab and the Reference Center for Special Immunobiologicals (Centro de Referência para Imunobiológicos Especiais - Crie) from Universidade Federal de São Paulo (Unifesp) was the first center designed for the municipality of São Paulo. In this context, the objectives of the present study were to describe the etiology of acute viral respiratory infection in children on prophylaxis with palivizumab and to analyze the rates of hospitalizations and deaths in this group of children.
Method This was a prospective study with infants who received palivizumab at the Crie-Unifesp in 2008. The project was approved by the Research Ethics Committee of the institution under n. 0342-08, and the signature of the informed consent by parents/guardians was required. The study included infants with indication of palivizumab according to the normative resolution SS n. 249, from July 13, 2007(15), that is, children younger than 1 year of age, born with gestational age lower than 29 weeks, after hospital discharge, and children younger than 2 years, with hemodynamically unstable cardiopathy or chronic pulmonary disease, who needed treatment in the 6 months prior to the period of the RSV season. Prophylaxis started in April 2008, including infants who received the first dose of palivizumab until the last month of June, in the dosage of 15mg/kg, via intramuscular (IM), in the vastus lateralis muscle of the thigh, with an interval of 30 days and maximum of five doses. The scheduling was performed grouping patients twice a week to share the flask of the immunobiological.
154
Data on the clinical records of the mother and the child and the reason for indication of the anti-RSV prophylaxis were collected on the day of the first dose of palivizumab by means of a questionnaire applied to the parents, besides the analysis of a medical form that indicated the administration of the monoclonal antibody. Patients were followed with monthly visits during all the season (April to September), and the parents were instructed to communicate the researcher in case the child presented acute respiratory tract infection during this period (passive vigilance). Furthermore, each patient was contacted by phone once a week to search for respiratory intercurrences and hospitalizations (active vigilance) during the entire study period. The acute respiratory tract infection was defined according to the World Health Organization(16) as the presence or not of fever, associated to running nose or congestion, sore throat, cough, sibilance, tachypnea or respiratory difficulty. In the presence of acute respiratory symptoms, a nasopharyngeal aspirate (NPA) was collected until the fourth day after the start of symptoms for the investigation of the following respiratory viruses: influenza A and B, parainfluenza 1, 2, and 3, adenovirus, RSV, rhinovirus, and metapneumovirus. For these patients, a second questionnaire was applied with clinical data on the current respiratory event. The RSV hospitalization rate was calculated using the rate of hospitalization for respiratory distress, multiplied by the number of children testing positive for RSV, divided by the number of children hospitalized for respiratory infection who underwent collection(13). The collection of the nasopharyngeal aspirate was conducted with probe n. 6 or 8, inserted in the nasopharynx after washing with saline solution at 0.9%. The aspirate was placed in a sterile flask with 5mL of saline solution at 0.9%. The sample was kept under refrigeration at 2-8°C and brought to the Laboratory of Virology of the institution in 2 hours maximum. The samples were fractionated, and an aliquot was centrifuged to perform the direct immunofluorescence and the other, was frozen at -80°C, for molecular testing. We performed direct immunofluorescence for influenza A and B, parainfluenza 1, 2 and 3, adenovirus, and RSV, according to the Light Diagnostics® Respiratory Screen Kit – Respiratory Panel 1 DFA (Chemicon International, Inc.). All samples underwent polymerase chain reaction preceded by reverse transcriptase for rhinovirus and human metapneumovirus(17-19). Numerical variables with normal distribution were compared by Student’s t test and those with abnormal distribution, by the Mann-Whitney test. Categorical variables were compared
Rev Paul Pediatr 2014;32(2):152-8.
Ana Isabel M. P. Monteiro et al
by chi-square or Fisher’s exact test. Statistical analysis were performed with the Statistical Package for the Social Sciences (SPSS), version 17, and significance was established at p
DOI: 10.1590/0103-0582201432214813
Respiratory infections in children up to two years of age on prophylaxis with palivizumab Infecções respiratórias em crianças menores de dois anos de idade submetidas a profilaxia com palivizumabe Infecciones respiratorias en niños menores de dos años de edad sometidos a profilaxis con palivizumabe Ana Isabel M. P. Monteiro1, Nancy Cristina J. Bellei1, Alessandra Ramos Sousa1, Amélia Miyashiro N. dos Santos1, Lily Yin Weckx1
ABSTRACT
Objective: To identify the viruses involved in acute respiratory tract infections and to analyze the rates of hospitalization and death in children on palivizumab prophylaxis. Methods: Prospective cohort of 198 infants up to one year old who were born before 29 weeks of gestational age and infants under two years old with hemodynamically unstable cardiopathy or chronic pulmonary disease who received prophylactic palivizumab against severe respiratory syncytial virus infections in 2008. During the study period, in each episode of acute respiratory tract infection, nasopharyngeal aspirate was collected to identify respiratory syncytial virus, adenovirus, parainfluenza 1, 2 and 3, influenza A and B by direct immunofluorescence, rhinovirus and metapneumovirus by polymerase chain reaction preceded by reverse transcription. Data regarding hospitalization and deaths were monitored. Results: Among the 198 studied infants, 117 (59.1%) presented acute respiratory tract infections, with a total of 175 episodes. Of the 76 nasopharyngeal aspirates collected during respiratory tract infections, 37 were positive, as follow: rhinovirus (75.7%), respiratory syncytial virus (18.9%), parainfluenza (8.1%), adenovirus 2 (2.7%), metapneumovirus (2.7%) and three samples presented multiple
Instituição: Escola Paulista de Medicina da Universidade Federal de São Paulo (Unifesp), São Paulo, SP, Brasil 1
Escola Paulista de Medicina da Unifesp, São Paulo, SP, Brasil
agents. Of the 198 children, 48 (24.4%) were hospitalized: 30 (15.2%) for non-infectious etiology and 18 (9.1%) for respiratory causes. Among these 18 children, one case of respiratory syncytial virus was identified. Two deaths were reported, but respiratory syncytial virus was not identified. Conclusions: During the prophylaxis period, low frequency of respiratory syncytial virus infections and low rates of hospitalization were observed, suggesting the benefit of palivizumab prophylaxis. Key-words: respiratory syncytial viruses; antibodies, monoclonal; respiratory tract infections/prevention & control; infant. RESUMO
Objetivo: Identificar os vírus envolvidos nos quadros de infecções agudas de trato respiratório e analisar as taxas de internação e de óbito em crianças submetidas à profilaxia com palivizumabe. Métodos: Coorte prospectiva com 198 crianças menores de um ano nascidas antes de 29 semanas de idade gestacional e crianças menores de dois anos com cardiopatia hemodinamicamente instável ou doença pulmonar crônica que receberam palivizumabe para profilaxia contra infecções graves pelo vírus
Endereço para correspondência: Ana Isabel M. P. Monteiro Rua Loefgreen, 1.998 CEP 04040-003 – São Paulo/SP E-mail: [email protected] Fonte Financiadora: Abbott Laboratories (Chicago, EUA) custeou os testes laboratoriais Conflito de interesse: Lily Yin Weckx realiza pesquisa clínica financiada pela GlaxoSmithKline, Merck Sharp & Dome, Novartis, Sanofi e Pfizer e palestras científicas para a GSK, MSD, Abbott, Sanofi, Novartis e Pfizer. Os demais autores não apresentam conflito de interesse Recebido em: 23/9/2013 Aprovado em: 27/11/2013
Rev Paul Pediatr 2014;32(2):152-8.
Ana Isabel M. P. Monteiro et al
sincicial respiratório, em 2008. No período do estudo, em cada episódio de infecção aguda do trato respiratório, coletou-se aspirado de nasofaringe para identificar vírus sincicial respiratório, adenovírus, parainfluenza 1, 2 e 3, influenza A e B por imunofluorescência direta, rinovírus e metapneumovírus por reação em cadeia de polimerase precedida de transcriptase reversa. Monitoraram-se internações e óbitos nesse grupo. Resultados: Das 198 crianças acompanhadas, 117 (59,1%) apresentaram infecções agudas de trato respiratório, totalizando 175 episódios. De 76 aspirados de nasofaringe coletados na vigência de infecções do trato respiratório, 37 foram positivos, encontrando-se: rinovírus (75,7%), vírus sincicial respiratório (18,9%), parainfluenza (8,1%), adenovírus (2,7%), metapneumovírus (2,7%) e múltiplos agentes em três amostras. Das 198 crianças, 48 (24,4%) foram internadas, sendo 30 (15,2%) por etiologia não respiratória e 18 (9,1%) por problemas respiratórios; entre os 18 casos, um foi por vírus sincicial respiratório. Duas crianças evoluíram para óbito, não tendo sido identificado o vírus sincicial respiratório. Conclusões: Na vigência de profilaxia, observou-se frequência baixa de infecções pelo vírus sincicial respiratório e baixo índice de hospitalizações, sugerindo benefício da profilaxia com palivizumabe. Palavras-chave: vírus sinciciais respiratórios; anticorpos monoclonais; infecções respiratórias/prevenção & controle; lactente. RESUMEN
Objetivo: Identificar los virus implicados en los cuadros de infecciones agudas de trato respiratorio y analizar las tasas de internación y de óbito en niños sometidos a la profilaxis con palivizumabe. Métodos: Cohorte prospectiva con 198 niños con menos de un año nacidas antes de 29 semanas de edad gestacional y niños con menos de dos años con cardiopatía hemodinámicamente inestable o enfermedad pulmonar crónica que recibieron palivizumabe para profilaxis contra infecciones graves por el virus sincitial respiratorio, en 2008. En el periodo de estudio, en cada episodio de infección aguda del trato respiratorio, se recogió aspirado de nasofaringe para identificar virus sincitial respiratorio, adenovirus, parainfluenza 1,2 y 3, influenza A y B por técnica del anticuerpo fluorescente directa, rinovirus y metapneumovirus por reacción en cadena de la polimerasa precedida por transcriptasa inversa. Se monitorean internaciones y óbitos en ese grupo.
Rev Paul Pediatr 2014;32(2):152-8.
Resultados: De los 198 niños seguidos, 117 (59,1%) presentaron infecciones agudas de trato respiratorio, totalizando 175 episodios. De 76 aspirados de nasofaringe recogidos en la vigencia de infecciones de trato respiratorio, 37 fueron positivos, encontrándose: rinovirus (75,7%), virus sincitial respiratorio (18,9%), parainfluenza (8,1%), adenovirus (2,7%), metapneumovirus (2,7%) y múltiples agentes en tres muestras. De los 198 niños, 48 (24,4%) fueron internados, siendo 30 (15,2%) por etiología no respiratoria y 18 (9,1%) por problemas respiratorios; entre los 18 casos, uno fue por virus sincitial respiratorio. Dos niños evolucionaron a óbito, no habiendo sido identificado el virus sincitial respiratorio. Conclusiones: En la vigencia de profilaxis, se observó frecuencia baja de infecciones por el virus sincitial respiratorio y bajo índice de hospitalizaciones, sugiriendo beneficio de la profilaxis con palivizumabe. Palabras clave: virus sincitiales respiratorios; anticuerpos monoclonales; infecciones respiratorias/ prevención y control; lactante.
Introduction Respiratory syncytial virus (RSV) is the major cause of acute lower respiratory tract infections in children younger than 2 years of age. It has a global distribution and a seasonal behavior. It is estimated that, worldwide, 33.8 million new episodes of acute lower respiratory tract infections by RSV occur annually in children under 5 years of age. Among these, about 3.4 million require hospitalization and 66 to 199 thousand children evolve to death, 99% occurring in developing countries(1). In general, primary RSV infection evolves as a common cold. However, approximately 25% of children under 2 years may present, in its first episode, infection of the lower airways, severe respiratory failure, need for ventilatory support, and, occasionally, death(2). Preterm children with chronic pulmonary disease, immunocompromised children, and children with heart diseases (particularly cyanotic) present greater risk of severe respiratory disease by RSV(3-6). Some environmental factors are also associated with higher incidence of RSV infections, such as exposure to smoke, attendance at day care centers, or household contact with other children(4,5). In tropical and subtropical countries, the occurrence of RSV infections is greater in fall and winter, with significant regional variations(7). Epidemiological studies on RSV in the Southeast region of Brazil showed that the infection by RSV starts in April, with peaks in May and June(2,8). More recent
153
Respiratory infections in children up to two years of age on prophylaxis with palivizumab
studies conducted in the municipality of São Paulo showed that the RSV season occurs from March to July, and it may start earlier or last up until August(9,10). There is no specific treatment for RSV infection(7,11), and prophylaxis with a specific anti-RSV monoclonal antibody (palivizumab) is widely recommended for high-risk cases(4-6). Its use demonstrated a reduction in the rates of hospitalization by RSV infection (55%), hospitalization in ICUs (57%), length of hospital stay (42%), and the number of days with need of oxygen by 40%(4,5,12,13). Palivizumab is, however, a high cost medication. In Brazil, until 2007, many children received this medication after judicial demands, not always based in technical criteria(14). The resolution SS n. 249, from July 13, 2007, from the Health Secretariat of the State of São Paulo(15), made palivizumab freely available for SUS (Brazilian public Unified Health System) patients who were classified as “highly recommended” by the Brazilian Pediatrics Society(5). Some centers were indicated by the State Secretariat as a place for the application of the palivizumab and the Reference Center for Special Immunobiologicals (Centro de Referência para Imunobiológicos Especiais - Crie) from Universidade Federal de São Paulo (Unifesp) was the first center designed for the municipality of São Paulo. In this context, the objectives of the present study were to describe the etiology of acute viral respiratory infection in children on prophylaxis with palivizumab and to analyze the rates of hospitalizations and deaths in this group of children.
Method This was a prospective study with infants who received palivizumab at the Crie-Unifesp in 2008. The project was approved by the Research Ethics Committee of the institution under n. 0342-08, and the signature of the informed consent by parents/guardians was required. The study included infants with indication of palivizumab according to the normative resolution SS n. 249, from July 13, 2007(15), that is, children younger than 1 year of age, born with gestational age lower than 29 weeks, after hospital discharge, and children younger than 2 years, with hemodynamically unstable cardiopathy or chronic pulmonary disease, who needed treatment in the 6 months prior to the period of the RSV season. Prophylaxis started in April 2008, including infants who received the first dose of palivizumab until the last month of June, in the dosage of 15mg/kg, via intramuscular (IM), in the vastus lateralis muscle of the thigh, with an interval of 30 days and maximum of five doses. The scheduling was performed grouping patients twice a week to share the flask of the immunobiological.
154
Data on the clinical records of the mother and the child and the reason for indication of the anti-RSV prophylaxis were collected on the day of the first dose of palivizumab by means of a questionnaire applied to the parents, besides the analysis of a medical form that indicated the administration of the monoclonal antibody. Patients were followed with monthly visits during all the season (April to September), and the parents were instructed to communicate the researcher in case the child presented acute respiratory tract infection during this period (passive vigilance). Furthermore, each patient was contacted by phone once a week to search for respiratory intercurrences and hospitalizations (active vigilance) during the entire study period. The acute respiratory tract infection was defined according to the World Health Organization(16) as the presence or not of fever, associated to running nose or congestion, sore throat, cough, sibilance, tachypnea or respiratory difficulty. In the presence of acute respiratory symptoms, a nasopharyngeal aspirate (NPA) was collected until the fourth day after the start of symptoms for the investigation of the following respiratory viruses: influenza A and B, parainfluenza 1, 2, and 3, adenovirus, RSV, rhinovirus, and metapneumovirus. For these patients, a second questionnaire was applied with clinical data on the current respiratory event. The RSV hospitalization rate was calculated using the rate of hospitalization for respiratory distress, multiplied by the number of children testing positive for RSV, divided by the number of children hospitalized for respiratory infection who underwent collection(13). The collection of the nasopharyngeal aspirate was conducted with probe n. 6 or 8, inserted in the nasopharynx after washing with saline solution at 0.9%. The aspirate was placed in a sterile flask with 5mL of saline solution at 0.9%. The sample was kept under refrigeration at 2-8°C and brought to the Laboratory of Virology of the institution in 2 hours maximum. The samples were fractionated, and an aliquot was centrifuged to perform the direct immunofluorescence and the other, was frozen at -80°C, for molecular testing. We performed direct immunofluorescence for influenza A and B, parainfluenza 1, 2 and 3, adenovirus, and RSV, according to the Light Diagnostics® Respiratory Screen Kit – Respiratory Panel 1 DFA (Chemicon International, Inc.). All samples underwent polymerase chain reaction preceded by reverse transcriptase for rhinovirus and human metapneumovirus(17-19). Numerical variables with normal distribution were compared by Student’s t test and those with abnormal distribution, by the Mann-Whitney test. Categorical variables were compared
Rev Paul Pediatr 2014;32(2):152-8.
Ana Isabel M. P. Monteiro et al
by chi-square or Fisher’s exact test. Statistical analysis were performed with the Statistical Package for the Social Sciences (SPSS), version 17, and significance was established at p
